A Journal on Biotechnology and Molecular Biology

Minerva Biotecnologica 2016 December;28(4):219-23

Substitution of conserved glycine with threonine in the active site of OsTrx23 by site-directed mutagenesis

Mitra ROODGAR NASHTA, Azar SHAHPIRI

Department of Biotechnology, College of Agriculture, Isfahan University of Technology, Isfahan, Iran

BACKGROUND: The main purpose of this study was to investigate the effect of an amino acid with side chain containing hydroxyl group in the active site on the activity of thioredoxins (Trxs).METHODS: A mutant protein with a threonine (Thr) substitution for the largely conserved glycine (Gly) at position 41 in the active site (WCGPC) of one of rice cytoplasmic Trx isoforms, OsTrx23, generated using site-directed mutagenesis and the activities of wild type (Wt) and mutant was compared using insulin assay under different pH.RESULTS: Insulin assay showed that whereas the activity of WtOsTrx23 were stable at different pH the activity of mutant G41TOsTrx23 was changed under different pH.CONCLUSIONS: The results indicate that the presence of Gly after second cysteine residue in the active site has an important role in stability of Trx under different pH conditions. The pH stability decreases considerably in the presence of a hydroxyl group in this position.